The aim of this proposal is to develop novel sets of inhibitors against the cell wall biosynthetic enzymes of M. tuberculosis. Many of these enzymes are responsible for the synthesis of the unique and essential components of Mycobacterium tuberculosis cell wall that are not found in the human host, making their biosynthesis an extremely attractive drug target. This proposal focuses on the implementation of a technologically advanced drug development program to design and synthesize novel inhibitors and advance screening hits. This project details a general medicinal chemistry hit development strategy to further advance hits from cycle ready targets in Projects 3 and will be applied to the pipeline enzyme targets as they are advanced to the refinement cycle. Thus methods outlined for drug discovery are designed to be complementary and are aimed to be applicable to all the enzyme targets. Screening hits from Project 3 will be analyzed to remove hits with undesirable functional groups and known frequent hitters. Hits and hit templates will then be identified and pdoritized for advancement based on relative activity both enzymatic and MIC against MDRTB as determined in Core A. The hits will then be advanced by elaboration using structure guided parallel synthesis, or by similarity searches of available data bases and virtual screening experiments. The goals is build up a strong structure activity relationship, and once high affinity inhibitors are identified with good activity against MDRTB, the emphasis will shift to optimizing the physical properties to maximize pharmacokinetics and bioactivity in vivo using the TB aerosol mouse infection model in close collaboration with Core B. At the completion of this process we aim to produce to leads suitable for clinical development.